Sci Rep. 2021 Feb 26;11(1):4787. doi: 10.1038/s41598-021-84114-y.

ABSTRACT

Ribosome biogenesis is tightly associated to plant metabolism due to the usage of ribosomes in the synthesis of proteins necessary to drive metabolic pathways. Given the central role of ribosome biogenesis in cell physiology, it is important to characterize the impact of different components involved in this process on plant metabolism. Double mutants of the Arabidopsis thaliana cytosolic 60S maturation factors REIL1 and REIL2 do not resume growth after shift to moderate 10 [Formula: see text] chilling conditions. To gain mechanistic insights into the metabolic effects of this ribosome biogenesis defect on metabolism, we developed TC-iReMet2, a constraint-based modelling approach that integrates relative metabolomics and transcriptomics time-course data to predict differential fluxes on a genome-scale level. We employed TC-iReMet2 with metabolomics and transcriptomics data from the Arabidopsis Columbia 0 wild type and the reil1-1 reil2-1 double mutant before and after cold shift. We identified reactions and pathways that are highly altered in a mutant relative to the wild type. These pathways include the Calvin-Benson cycle, photorespiration, gluconeogenesis, and glycolysis. Our findings also indicated differential NAD(P)/NAD(P)H ratios after cold shift. TC-iReMet2 allows for mechanistic hypothesis generation and interpretation of system biology experiments related to metabolic fluxes on a genome-scale level.

PMID:33637852 | DOI:10.1038/s41598-021-84114-y

Cancer Immunol Res. 2021 Feb 26:canimm.0637.2020. doi: 10.1158/2326-6066.CIR-20-0637. Online ahead of print.

ABSTRACT

The theory of cancer immunoediting, which describes the dynamic interactions between tumors and host immune cells that shape the character of each compartment, is foundational for understanding cancer immunotherapy. Few models exist that facilitate in-depth study of each of the three canonical phases of immunoediting: elimination, equilibrium, and escape. Here, we utilized NPK-C1, a transplantable prostate tumor model that we found recapitulated the three phases of immunoediting spontaneously in immunocompetent animals. Given that a significant portion of NPK-C1 tumors reliably progressed to the escape phase, we were able to delineate cell types and mechanisms differentially prevalent in equilibrium versus escape phases. Using high-dimensional flow cytometry, we found that activated CD4+ effector T cells were enriched in regressing tumors, highlighting a role for CD4+ T cells in antitumor immunity. CD8+ T cells were also important for NPK-C1 control; specifically, central memory-like cytotoxic CD8+ T cells. Regulatory T cells (Tregs), as a whole, were counterintuitively enriched in regressing tumors; however, high-dimensional analysis revealed their significant phenotypic diversity, with a number of Treg subpopulations enriched in progressing tumors. In the myeloid compartment, we found that iNOS+ DC-like cells are enriched in regressing tumors, whereas CD103+ DCs were associated with late-stage tumor progression. In total, these analyses of the NPK-C1 model provide novel insights into the roles of lymphoid and myeloid populations throughout the cancer immunoediting process and highlight a role for multi-dimensional, flow-based analyses to more deeply understand immune cell dynamics in the tumor microenvironment.

PMID:33637604 | DOI:10.1158/2326-6066.CIR-20-0637

Sci Immunol. 2021 Feb 26;6(56):eabb7221. doi: 10.1126/sciimmunol.abb7221.

ABSTRACT

Mast cells (MCs) play a pathobiologic role in type 2 (T2) allergic inflammatory diseases of the airway, including asthma and chronic rhinosinusitis with nasal polyposis (CRSwNP). Distinct MC subsets infiltrate the airway mucosa in T2 disease, including subepithelial MCs expressing the proteases tryptase and chymase (MCTC) and epithelial MCs expressing tryptase without chymase (MCT). However, mechanisms underlying MC expansion and the transcriptional programs underlying their heterogeneity are poorly understood. Here, we use flow cytometry and single-cell RNA-sequencing (scRNA-seq) to conduct a comprehensive analysis of human MC hyperplasia in CRSwNP, a T2 cytokine-mediated inflammatory disease. We link discrete cell surface phenotypes to the distinct transcriptomes of CRSwNP MCT and MCTC, which represent polarized ends of a transcriptional gradient of nasal polyp MCs. We find a subepithelial population of CD38highCD117high MCs that is markedly expanded during T2 inflammation. These CD38highCD117high MCs exhibit an intermediate phenotype relative to the expanded MCT and MCTC subsets. CD38highCD117high MCs are distinct from circulating MC progenitors and are enriched for proliferation, which is markedly increased in CRSwNP patients with aspirin-exacerbated respiratory disease, a severe disease subset characterized by increased MC burden and elevated MC activation. We observe that MCs expressing a polyp MCT-like effector program are also found within the lung during fibrotic diseases and asthma, and further identify marked differences between MCTC in nasal polyps and skin. These results indicate that MCs display distinct inflammation-associated effector programs and suggest that in situ MC proliferation is a major component of MC hyperplasia in human T2 inflammation.

PMID:33637594 | DOI:10.1126/sciimmunol.abb7221

Sci Adv. 2021 Feb 26;7(9):eabe7999. doi: 10.1126/sciadv.abe7999. Print 2021 Feb.

ABSTRACT

Eukaryotic cells rely on endocytosis to regulate their plasma membrane proteome and lipidome. Most eukaryotic groups, except fungi and animals, have retained the evolutionary ancient TSET complex as an endocytic regulator. Unlike other coatomer complexes, structural insight into TSET is lacking. Here, we reveal the molecular architecture of plant TSET [TPLATE complex (TPC)] using an integrative structural approach. We identify crucial roles for specific TSET subunits in complex assembly and membrane interaction. Our data therefore generate fresh insight into the differences between the hexameric TSET in Dictyostelium and the octameric TPC in plants. Structural elucidation of this ancient adaptor complex represents the missing piece in the coatomer puzzle and vastly advances our functional as well as evolutionary insight into the process of endocytosis.

PMID:33637534 | DOI:10.1126/sciadv.abe7999

Sci Adv. 2021 Feb 26;7(9):eabb7921. doi: 10.1126/sciadv.abb7921. Print 2021 Feb.

ABSTRACT

Bioactive metabolites have wide-ranging biological activities and are a potential source of future research and therapeutic tools. Here, we use nanovibrational stimulation to induce osteogenic differentiation of mesenchymal stem cells, in the absence of off-target, nonosteogenic differentiation. We show that this differentiation method, which does not rely on the addition of exogenous growth factors to culture media, provides an artifact-free approach to identifying bioactive metabolites that specifically and potently induce osteogenesis. We first identify a highly specific metabolite, cholesterol sulfate, an endogenous steroid. Next, a screen of other small molecules with a similar steroid scaffold identified fludrocortisone acetate with both specific and highly potent osteogenic-inducing activity. Further, we implicate cytoskeletal contractility as a measure of osteogenic potency and cell stiffness as a measure of specificity. These findings demonstrate that physical principles can be used to identify bioactive metabolites and then enable optimization of metabolite potency can be optimized by examining structure-function relationships.

PMID:33637520 | DOI:10.1126/sciadv.abb7921

Kidney Int. 2021 Mar;99(3):498-510. doi: 10.1016/j.kint.2020.08.039.

ABSTRACT

Chronic kidney disease (CKD) and acute kidney injury (AKI) are common, heterogeneous, and morbid diseases. Mechanistic characterization of CKD and AKI in patients may facilitate a precision-medicine approach to prevention, diagnosis, and treatment. The Kidney Precision Medicine Project aims to ethically and safely obtain kidney biopsies from participants with CKD or AKI, create a reference kidney atlas, and characterize disease subgroups to stratify patients based on molecular features of disease, clinical characteristics, and associated outcomes. An additional aim is to identify critical cells, pathways, and targets for novel therapies and preventive strategies. This project is a multicenter prospective cohort study of adults with CKD or AKI who undergo a protocol kidney biopsy for research purposes. This investigation focuses on kidney diseases that are most prevalent and therefore substantially burden the public health, including CKD attributed to diabetes or hypertension and AKI attributed to ischemic and toxic injuries. Reference kidney tissues (for example, living-donor kidney biopsies) will also be evaluated. Traditional and digital pathology will be combined with transcriptomic, proteomic, and metabolomic analysis of the kidney tissue as well as deep clinical phenotyping for supervised and unsupervised subgroup analysis and systems biology analysis. Participants will be followed prospectively for 10 years to ascertain clinical outcomes. Cell types, locations, and functions will be characterized in health and disease in an open, searchable, online kidney tissue atlas. All data from the Kidney Precision Medicine Project will be made readily available for broad use by scientists, clinicians, and patients.

PMID:33637194 | DOI:10.1016/j.kint.2020.08.039

Cell. 2021 Feb 17:S0092-8674(21)00092-1. doi: 10.1016/j.cell.2021.01.049. Online ahead of print.

ABSTRACT

Clonal hematopoiesis, a condition in which individual hematopoietic stem cell clones generate a disproportionate fraction of blood leukocytes, correlates with higher risk for cardiovascular disease. The mechanisms behind this association are incompletely understood. Here, we show that hematopoietic stem cell division rates are increased in mice and humans with atherosclerosis. Mathematical analysis demonstrates that increased stem cell proliferation expedites somatic evolution and expansion of clones with driver mutations. The experimentally determined division rate elevation in atherosclerosis patients is sufficient to produce a 3.5-fold increased risk of clonal hematopoiesis by age 70. We confirm the accuracy of our theoretical framework in mouse models of atherosclerosis and sleep fragmentation by showing that expansion of competitively transplanted Tet2-/- cells is accelerated under conditions of chronically elevated hematopoietic activity. Hence, increased hematopoietic stem cell proliferation is an important factor contributing to the association between cardiovascular disease and clonal hematopoiesis.

PMID:33636128 | DOI:10.1016/j.cell.2021.01.049

Cell. 2021 Feb 9:S0092-8674(21)00158-6. doi: 10.1016/j.cell.2021.02.008. Online ahead of print.

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of the ongoing coronavirus disease 2019 (COVID-19) pandemic. Understanding of the RNA virus and its interactions with host proteins could improve therapeutic interventions for COVID-19. By using icSHAPE, we determined the structural landscape of SARS-CoV-2 RNA in infected human cells and from refolded RNAs, as well as the regulatory untranslated regions of SARS-CoV-2 and six other coronaviruses. We validated several structural elements predicted in silico and discovered structural features that affect the translation and abundance of subgenomic viral RNAs in cells. The structural data informed a deep-learning tool to predict 42 host proteins that bind to SARS-CoV-2 RNA. Strikingly, antisense oligonucleotides targeting the structural elements and FDA-approved drugs inhibiting the SARS-CoV-2 RNA binding proteins dramatically reduced SARS-CoV-2 infection in cells derived from human liver and lung tumors. Our findings thus shed light on coronavirus and reveal multiple candidate therapeutics for COVID-19 treatment.

PMID:33636127 | DOI:10.1016/j.cell.2021.02.008

Kaohsiung J Med Sci. 2021 Feb 26. doi: 10.1002/kjm2.12375. Online ahead of print.

ABSTRACT

Immunogenic cell death (ICD) refers to a form of regulated cell death that activates adaptive immunity, forming long-term immunological memory. Using chemotherapeutic drugs to induce ICD in cancer cells can help create an inflamed, immunogenic tumor environment, key for optimal immune checkpoint blockade (ICB) therapy response. ICB targets immune checkpoints such as cytotoxic T-lymphocyte-associated antigen 4 (CTLA4) and programmed death 1 (PD-1). Durable responses and better quality of life in ICB patients compared with many other treatments has prompted additional investigation into its therapeutic potential and possible approaches, in an effort to further understand the functions of the costimulatory molecules and how new treatments may be designed. In this review, we will summarize ICD induction, including stress responses, damage-associated molecular patterns, and various assays by which immunogenicity is evaluated in dying cells. In addition, the mechanisms and biomarkers underlying the CTLA4 and PD-1 pathways of checkpoint blockade will be covered. Finally, we will review the synergistic effects of ICD induction combined with ICB therapy, as well as combination blockade therapies involving the use of multiple drugs.

PMID:33636043 | DOI:10.1002/kjm2.12375

PLoS Pathog. 2021 Feb 26;17(2):e1009325. doi: 10.1371/journal.ppat.1009325. eCollection 2021 Feb.

ABSTRACT

Apicomplexan parasites are causative agents of major human diseases. Calcium Dependent Protein Kinases (CDPKs) are crucial components for the intracellular development of apicomplexan parasites and are thus considered attractive drug targets. CDPK7 is an atypical member of this family, which initial characterization suggested to be critical for intracellular development of both Apicomplexa Plasmodium falciparum and Toxoplasma gondii. However, the mechanisms via which it regulates parasite replication have remained unknown. We performed quantitative phosphoproteomics of T. gondii lacking TgCDPK7 to identify its parasitic targets. Our analysis lead to the identification of several putative TgCDPK7 substrates implicated in critical processes like phospholipid (PL) synthesis and vesicular trafficking. Strikingly, phosphorylation of TgRab11a via TgCDPK7 was critical for parasite intracellular development and protein trafficking. Lipidomic analysis combined with biochemical and cellular studies confirmed that TgCDPK7 regulates phosphatidylethanolamine (PE) levels in T. gondii. These studies provide novel insights into the regulation of these processes that are critical for parasite development by TgCDPK7.

PMID:33635921 | DOI:10.1371/journal.ppat.1009325

Clin Transl Med. 2021 Feb;11(2):e307. doi: 10.1002/ctm2.307.

ABSTRACT

BACKGROUND: Hepatocellular carcinoma (HCC) is a malignant disease with high morbidity and mortality, and the molecular mechanism for the genesis and progression is complex and heterogeneous. Biomarker discovery is crucial for the personalized and precision treatment of HCC. The accumulation of reported microRNA biomarkers makes it possible to combine computational identification with experimental validation to accelerate the discovery of novel biomarker.

RESULTS: In the present work, we applied a rational computer-aided biomarker discovery model to screen for the HCC diagnosis biomarker. Two HCC-associated networks were constructed based on the microRNA and mRNA expression profiles, and the potential microRNA biomarkers were identified based on their unique regulatory and influential power in the network. These putative biomarkers were then experimentally validated. One prominent example among these identified biomarkers is MiR-378a-3p: It was shown to independently regulate several important transcription factors such as PLAGL2 and β-catenin, affecting the β-catenin signaling. Such mechanism may indicate a potential tumor suppressor role of MiR-378a-3p and the impact of its abnormal expression on the cell growth and invasion of HCC.

CONCLUSIONS: A bioinformatics model with network topological and functional characterization was successfully applied to the identification of HCC biomarkers. The predicted microRNA biomarkers were than validated with experiments using human HCC cell lines, model animal, and clinical specimens. The results confirmed the prediction by our proposed model that miR-378a-3p was a putative biomarker for diagnosis and prognosis of HCC.

PMID:33634974 | DOI:10.1002/ctm2.307

Elife. 2021 Feb 26;10:e64000. doi: 10.7554/eLife.64000. Online ahead of print.

ABSTRACT

Automated visual tracking of animals is rapidly becoming an indispensable tool for the study of behavior. It offers a quantitative methodology by which organisms' sensing and decision-making can be studied in a wide range of ecological contexts. Despite this, existing solutions tend to be challenging to deploy in practice, especially when considering long and/or high-resolution video-streams. Here, we present TRex, a fast and easy-to-use solution for tracking a large number of individuals simultaneously using background-subtraction with real-time (60Hz) tracking performance for up to approximately 256 individuals and estimates 2D visual-fields, outlines, and head/rear of bilateral animals, both in open and closed-loop contexts. Additionally, TRex offers highly-accurate, deep-learning-based visual identification of up to approximately 100 unmarked individuals, where it is between 2.5-46.7 times faster, and requires 2-10 times less memory, than comparable software (with relative performance increasing for more organisms/longer videos) and provides interactive data-exploration within an intuitive, platform-independent graphical user-interface.

PMID:33634789 | DOI:10.7554/eLife.64000

Autophagy. 2021 Feb 8:1-382. doi: 10.1080/15548627.2020.1797280. Online ahead of print.

ABSTRACT

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.

PMID:33634751 | DOI:10.1080/15548627.2020.1797280

Interface Focus. 2021 Apr 6;11(2):20200043. doi: 10.1098/rsfs.2020.0043. Epub 2021 Feb 12.

ABSTRACT

Many insects can detect carbon dioxide (CO2) plumes using a conserved receptor made up of members of the gustatory receptor (Gr) family Gr1, Gr2 and Gr3. Mosquitoes are attracted to host animals for blood meals using plumes of CO2 in the exhaled breath using the receptor expressed in the A neuron of the capitate peg sensilla type on the maxillary palps. The receptor is known to also detect several other classes of odorants, including ones emitted from human skin. Here, we discover that a common skin odorant, butyric acid, can cause a phasic activation followed by an unusually prolonged tonic activity after the stimulus is over in the CO2 neurons of mosquitoes. The effect is conserved in both Aedes aegypti and Anopheles gambiae mosquitoes. This raises a question about its role in a mosquito's preference for the skin odour of different individuals. Butyric acid belongs to a small number of odorants known to cause the prolonged activation of the CO2 receptor. A chemical informatic analysis identifies a specific set of physico-chemical features that can be used in a machine learning predictive model for the prolonged activators. Interestingly, this set is different from physico-chemical features selected for activators or inhibitors, indicating that each has a distinct structural basis. The structural understanding opens up an opportunity to find novel ligands to manipulate the CO2 receptor and mosquito behaviour.

PMID:33633836 | PMC:PMC7898149 | DOI:10.1098/rsfs.2020.0043

Front Genet. 2021 Feb 9;12:617059. doi: 10.3389/fgene.2021.617059. eCollection 2021.

ABSTRACT

Current methods of high-throughput molecular and genomic analyses enabled to reconstruct thousands of human molecular pathways. Knowledge of molecular pathways structure and architecture taken along with the gene expression data can help interrogating the pathway activation levels (PALs) using different bioinformatic algorithms. In turn, the pathway activation profiles can characterize molecular processes, which are differentially regulated and give numeric characteristics of the extent of their activation or inhibition. However, different pathway nodes may have different functions toward overall pathway regulation, and calculation of PAL requires knowledge of molecular function of every node in the pathway in terms of its activator or inhibitory role. Thus, high-throughput annotation of functional roles of pathway nodes is required for the comprehensive analysis of the pathway activation profiles. We proposed an algorithm that identifies functional roles of the pathway components and applied it to annotate 3,044 human molecular pathways extracted from the Biocarta, Reactome, KEGG, Qiagen Pathway Central, NCI, and HumanCYC databases and including 9,022 gene products. The resulting knowledgebase can be applied for the direct calculation of the PALs and establishing large scale profiles of the signaling, metabolic, and DNA repair pathway regulation using high throughput gene expression data. We also provide a bioinformatic tool for PAL data calculations using the current pathway knowledgebase.

PMID:33633781 | PMC:PMC7900570 | DOI:10.3389/fgene.2021.617059

Sci Rep. 2021 Feb 25;11(1):4577. doi: 10.1038/s41598-021-83883-w.

ABSTRACT

Idiopathic forms of Focal Segmental Glomerulosclerosis (FSGS) are caused by circulating permeability factors, which can lead to early recurrence of FSGS and kidney failure after kidney transplantation. In the past three decades, many research endeavors were undertaken to identify these unknown factors. Even though some potential candidates have been recently discussed in the literature, "the" actual factor remains elusive. Therefore, there is an increased demand in FSGS research for the use of novel technologies that allow us to study FSGS from a yet unexplored angle. Here, we report the successful treatment of recurrent FSGS in a patient after living-related kidney transplantation by removal of circulating factors with CytoSorb apheresis. Interestingly, the classical published circulating factors were all in normal range in this patient but early disease recurrence in the transplant kidney and immediate response to CytoSorb apheresis were still suggestive for pathogenic circulating factors. To proof the functional effects of the patient's serum on podocytes and the glomerular filtration barrier we used a podocyte cell culture model and a proteinuria model in zebrafish to detect pathogenic effects on the podocytes actin cytoskeleton inducing a functional phenotype and podocyte effacement. We then performed Raman spectroscopy in the < 50 kDa serum fraction, on cultured podocytes treated with the FSGS serum and in kidney biopsies of the same patient at the time of transplantation and at the time of disease recurrence. The analysis revealed changes in podocyte metabolome induced by the FSGS serum as well as in focal glomerular and parietal epithelial cell regions in the FSGS biopsy. Several altered Raman spectra were identified in the fractionated serum and metabolome analysis by mass spectrometry detected lipid profiles in the FSGS serum, which were supported by disturbances in the Raman spectra. Our novel innovative analysis reveals changed lipid metabolome profiles associated with idiopathic FSGS that might reflect a new subtype of the disease.

PMID:33633212 | DOI:10.1038/s41598-021-83883-w

Science. 2021 Feb 25:eabf7117. doi: 10.1126/science.abf7117. Online ahead of print.

ABSTRACT

Long-read and strand-specific sequencing technologies together facilitate the de novo assembly of high-quality haplotype-resolved human genomes without parent-child trio data. We present 64 assembled haplotypes from 32 diverse human genomes. These highly contiguous haplotype assemblies (average contig N50: 26 Mbp) integrate all forms of genetic variation even across complex loci. We identify 107,590 structural variants (SVs), of which 68% are not discovered by short-read sequencing, and 278 SV hotspots (spanning megabases of gene-rich sequence). We characterize 130 of the most active mobile element source elements and find that 63% of all SVs arise by homology-mediated mechanisms. This resource enables reliable graph-based genotyping from short reads of up to 50,340 SVs, resulting in the identification of 1,526 expression quantitative trait loci as well as SV candidates for adaptive selection within the human population.

PMID:33632895 | DOI:10.1126/science.abf7117

J Neurosci. 2021 Feb 25:JN-RM-2593-19. doi: 10.1523/JNEUROSCI.2593-19.2021. Online ahead of print.

ABSTRACT

Common fusion machinery mediates the Ca2+-dependent exocytosis of synaptic vesicles (SVs) and dense-core vesicles (DCVs). Previously, Synapsin Ia (Syn Ia) was found to localize to SVs, essential for mobilizing SVs to the plasma membrane through phosphorylation. However, whether (or how) the phosphoprotein Syn Ia plays a role in regulating DCV exocytosis remains unknown. To answer these questions, we measured the dynamics of DCV exocytosis by using single-vesicle amperometry in PC12 cells (derived from the pheochromocytoma of rats of unknown sex) overexpressing wild-type or phosphodeficient Syn Ia. We found that overexpression of phosphodeficient Syn Ia decreased the DCV secretion rate, specifically via residues previously shown to undergo calmodulin-dependent kinase (CaMK)-mediated phosphorylation (S9, S566, and S603). Moreover, the fusion pore kinetics during DCV exocytosis were found to be differentially regulated by Syn Ia and two phosphodeficient Syn Ia mutants (Syn Ia-S62A and Syn Ia-S9,566,603A). Kinetic analysis suggested that Syn Ia may regulate the closure and dilation of DCV fusion pores via these sites, implying the potential interactions of Syn Ia with certain DCV proteins involved in the regulation of fusion pore dynamics. Furthermore, we predicted the interaction of Syn Ia with several DCV proteins, including Synaptophysin (Syp) and soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins. By immunoprecipitation, we found that Syn Ia interacted with Syp via phosphorylation. Moreover, a proximity ligation assay (PLA) confirmed their phosphorylation-dependent, in situ interaction on DCVs. Together, these findings reveal a phosphorylation-mediated regulation of DCV exocytosis by Syn Ia.Significance StatementAlthough they exhibit distinct exocytosis dynamics upon stimulation, synaptic vesicles (SVs) and dense-core vesicles (DCVs) may undergo co-release in neurons and neuroendocrine cells through an undefined molecular mechanism. Synapsin Ia (Syn Ia) is known to recruit SVs to the plasma membrane via phosphorylation. Here, we examined whether Syn Ia also affects the dynamics of DCV exocytosis. We showed that Syn Ia regulates the DCV secretion rate and fusion pore kinetics during DCV exocytosis. Moreover, Syn Ia-mediated regulation of DCV exocytosis depends on phosphorylation. We further found that Syn Ia interacts with Synaptophysin on DCVs in a phosphorylation-dependent manner. Thus, these results suggest that Syn Ia may regulate the release of DCVs via phosphorylation.

PMID:33632727 | DOI:10.1523/JNEUROSCI.2593-19.2021

J Nanobiotechnology. 2021 Feb 25;19(1):59. doi: 10.1186/s12951-021-00806-7.

ABSTRACT

Virus-like particles (VLPs) are virus-derived structures made up of one or more different molecules with the ability to self-assemble, mimicking the form and size of a virus particle but lacking the genetic material so they are not capable of infecting the host cell. Expression and self-assembly of the viral structural proteins can take place in various living or cell-free expression systems after which the viral structures can be assembled and reconstructed. VLPs are gaining in popularity in the field of preventive medicine and to date, a wide range of VLP-based candidate vaccines have been developed for immunization against various infectious agents, the latest of which is the vaccine against SARS-CoV-2, the efficacy of which is being evaluated. VLPs are highly immunogenic and are able to elicit both the antibody- and cell-mediated immune responses by pathways different from those elicited by conventional inactivated viral vaccines. However, there are still many challenges to this surface display system that need to be addressed in the future. VLPs that are classified as subunit vaccines are subdivided into enveloped and non- enveloped subtypes both of which are discussed in this review article. VLPs have also recently received attention for their successful applications in targeted drug delivery and for use in gene therapy. The development of more effective and targeted forms of VLP by modification of the surface of the particles in such a way that they can be introduced into specific cells or tissues or increase their half-life in the host is likely to expand their use in the future. Recent advances in the production and fabrication of VLPs including the exploration of different types of expression systems for their development, as well as their applications as vaccines in the prevention of infectious diseases and cancers resulting from their interaction with, and mechanism of activation of, the humoral and cellular immune systems are discussed in this review.

PMID:33632278 | DOI:10.1186/s12951-021-00806-7

Plant Physiol. 2021 Feb 25;185(1):228-239. doi: 10.1093/plphys/kiaa019.

ABSTRACT

Optimal plant growth performance requires that the presence and action of growth signals, such as gibberellins (GAs), are coordinated with the availability of photo-assimilates. Here, we studied the links between GA biosynthesis and carbon availability, and the subsequent effects on growth. We established that carbon availability, light and dark cues, and the circadian clock ensure the timing and magnitude of GA biosynthesis and that disruption of these factors results in reduced GA levels and expression of downstream genes. Carbon-dependent nighttime induction of gibberellin 3-beta-dioxygenase 1 (GA3ox1) was severely hampered when preceded by reduced daytime light availability, leading specifically to reduced bioactive GA4 levels, and coinciding with a decline in leaf expansion rate during the night. We attributed this decline in leaf expansion mostly to reduced photo-assimilates. However, plants in which GA limitation was alleviated had significantly improved leaf expansion, demonstrating the relevance of GAs in growth control under varying carbon availability. Carbon-dependent expression of upstream GA biosynthesis genes (Kaurene synthase and gibberellin 20 oxidase 1, GA20ox1) was not translated into metabolite changes within this short timeframe. We propose a model in which the extent of nighttime biosynthesis of bioactive GA4 by GA3ox1 is determined by nighttime consumption of starch reserves, thus providing day-to-day adjustments of GA responses.

PMID:33631808 | DOI:10.1093/plphys/kiaa019